We have successfully induced leukemic cells from myelogenous leukemia patients and HL-60 promyelocyte cell line to differentiate and mature to macrophages in a liquid culture assay. Conditioned culture medium (CM) of PHA activated normal T cells contains the maturation inducing activity. Analysis of total DNA content of single cells by flow cytometry demonstrated clearly that differentiated macrophages possess the same aneuploid DNA stemline as the undifferentiated patient cells, indicating leukemic cells can differentiate independently of genetic abnormality. The developmental process revealed that CM first induced most of the leukemic cells for one cell cycle division, then suppressed the DNA replication preceding mature cell expression. This process has also been delineated by reduction of RNA content, development of adherent cells, morphology and acquisition of membrane receptors and functions. We hope these analyses of proliferation and maturation will represent a new and practical method to study leukemia. We are proposing to study leukemic patient cells with simultaneous application of the methods probing proliferation and differentiation defects. This includes the investigation of a lack of the capacity of stop leukemic cells from replicating, by comparison of the suppressive activity of CM of patients with normal on the proliferation of HL-60 cells. Microaasay technique to quantitate replicating cell number of autoradiography will be used. A possible excess or lack of proliferative stimulation will be studied by cross-testing CM of patients and of normal controls with patient and normal cells. Similar methods will be u sed with the culture assay to analyze whether there is a lack of CM maturation inducing activity for their own cells, thereby blocking their differentiation. The development of isolated immature cells of normal bone marrow could be compared. We propose also to identify the subset of T cells producing the activities for cells proliferation, differentiation and suppression of p roliferation. Combining assays of proliferation and differantiation on the same cells will enable us to understand the controls of the leukemic cell growth.